Effects of natural compounds on the accuracy of 8-oxo-7,8-dihydro-2'-deoxyguanosine translesion synthesis

Translesion synthesis (TLS) is a DNA damage tolerance mechanism that relies on a series of specialized DNA polymerases able to bypass a lesion on a DNA template strand during replication or post-repair synthesis. Specialized TLS DNA polymerases pursue replication by inserting a base opposite to this lesion, correctly or incorrectly depending on the lesion nature, involved DNA polymerase(s), sequence context and still unknown factors.

To measure the correct or mutagenic outcome of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) bypass by TLS, a primer-extension assay was performed in vitro on a template DNA bearing this lesion in the presence of nuclear proteins extracted from human intestinal epithelial cells (FHs 74 Int cell line); the reaction products were analysed by both denaturing capillary electrophoresis (to measure the yield of translesion elongation) and pyrosequencing (to determine the identity of the nucleotide inserted in front of the lesion). The influence of several natural compounds on the correct or mutagenic outcome of TLS through 8-oxodG was then evaluated, in two experimental conditions, by adding the polyphenol either (i) to the reaction mix during the primer extension assay; or (ii) to the culture medium, 24h before cell harvest and nuclear proteins extraction. Some of the tested compounds significantly influenced the outcome of translesion synthesis, either through an error-free or a mutagenic pathway.

Publication History

Article published online:
13 December 2021